Differentiated cell cultures are being used with increasing frequency in biomedical research. An unknown but significant number of these will become infected with mycoplasmas, rendering them useless for controlled studies. Most of the conclusions and indirect assay systems for mycoplasma infection of cell cultures are based on data from fibroblast systems. Data are available that indicate that some of these conclusions do not apply to other differentiated cell cultures. The objective of this proposal is to perform controlled prospective studies on fibroblast, lymphocyte, urinary epithelium, alveolar epithelium and teratocarcinoma cell cultures that are separately infected with a variety of mycoplasmal species. Parameters to be studied include a) incidence of mycoplasmal infection of primary cultures established from tissues that may contain mycoplasmas; b) long term effect of infection on cultures; c) effect on growth rate of cells; d) concentration of mycoplasmas in culture supernatants; e) degree of cytadsorption of mycoplasmas; f) effectiveness of four assay techniques to detect mycoplasmas in situ and in an indicator cell line; g) methods to prevent infection of primary cell; and h) studies of interest in specific differentiated cell cultures. Studies will also be performed to determine if alveolar epithelial and urinary epithelial cell cultures might be models of pathogenic mycoplasmas that infect these sites. Results of these studies will have practical and immediate significance in all in vitro studies that utilize differentiated cell cultures.